The competing renewal of this revised Program Project grant is organized around a unifying research theme, the pathogenesis and prevention of periventricular leukomalacia (PVL), the major form of brain injury in the premature infant. The proposed research is based primarily on the Program's many scientific discoveries of the previous cycle and on exciting preliminary data. The overall Program is composed of four integrated projects, one in clinical neuroscience and three in basic neuroscience. The first Project addresses the cellular and molecular pathology of PVL in human brain. Work hi the first cycle defined the key cellular target in PVL to be the pre-myelinating oligodendrocyte, i.e., pre-oligo, the death of which was shown to be associated with marked microgliosis and reactive astrocytosis and evidence for attack by reactive oxygen and nitrogen species (ROS/RNS). In the current proposal the degree of pre-oligo loss will be quantitated; the cellular origins of RNS identified; the role of microglia and astrocytes in ROS/RNS toxicity determined; and the extent and nature of neuronal/axonal impairment delineated. The second Project will address the cellular and molecular pathways of oxidative injury to pre-oligos, with an emphasis on microglia and RNS attack. Areas of focus include delineation of the mechanisms of injury to pre-oligos triggered by microglia, the molecular pathways responsible for RNS toxicity, and the role for RNS toxicity in in vivo white matter (WM) injury caused by hypoxia-ischemia or infection/inflammation or both insults. The third Project addresses the role of excitotoxicity in the pathogenesis of PVL. Emphases of the project are the expression of GluRs in normal human WM and in PVL, the toxic mechanisms downstream from ionotropic glutamate receptor (GluR) activation that are intrinsic to the pre-oligo or that involve other glial cell types, the sensitizing effect of subthreshold hypoxia on the impact of a subsequent insult, and the role of intracellular Ca2+and mitochondrial dysfunction in pre-oligo excitotoxicity. The fourth Project addresses the roles of exogenous and endogenous activators of microglia and of innate immunity in the pathogenesis of PVL. Research emphases include delineation of heat-shock protein 60 (HSP60) as a key endogenous activator of microglia and innate immunity and secondarily, thereby, injury to pre-oligos, the involvement of HSP60 in hypoxic-ischemic WM injury in the developing rodent and in human PVL, and the expression of key Toll-like receptors in developing human WM and in PVL. The progress of the overall Program in the first cycle and the proposed work building on that progress should lead to insights that allow formulation of preventative interventions suitable for clinical trial.